Impact of Phthalates on the Male: Frog and Rabbit Models

EPA Grant Number: R829429
Title: Impact of Phthalates on the Male: Frog and Rabbit Models
Investigators: Veeramachaneni, D. N. Rao
Institution: Colorado State University
EPA Project Officer: Klieforth, Barbara I
Project Period: March 1, 2002 through February 28, 2005 (Extended to February 28, 2007)
Project Amount: $852,709
RFA: Children's Vulnerability to Toxic Substances in the Environment (2001) RFA Text |  Recipients Lists
Research Category: Children's Health , Human Health , Environmental Justice


Disturbingly high levels of phthalates have been found in women of reproductive age (Environ Health Perspect 108:979, 2000). Although in utero and/or infantile exposures to pollutants such as phthalates have been associated with increasing incidence of reproductive disorders, substantiating data, particularly at environmentally relevant levels, are scant.

We hypothesize that exposure to phthalates, even at relatively low concentrations, during differentiation of the reproductive system alters reproductive function as adults. We propose to test this hypothesis in two animal models, an amphibian, Xenopus laevis, and a non-rodent mammal, the rabbit. The former facilitates transdermal exposure and evaluation of an easy-to-monitor, unique thyroid hormone-dependent event, the metamorphosis, while the latter facilitates longitudinal evaluations of hormones, semen parameters and sexual capacity. Furthermore, rabbits, unlike rodents, have a relatively long infantile period of reproductive development more closely approximating the situation in humans. Dermal route of exposure is particularly pertinent to children's vulnerability to toxic substances in the environment, since children have a greater ratio of surface area to body weight than adults.


Tadpoles will be exposed to 0 (vehicle-only control), 0.1, 0.5, 1, or 5 ppm dibutyl phthalate between 3 and 12 wk of life (beginning of sexual differentiation of gonads through completion of metamorphosis). Rabbits will be exposed in utero and through lactation. Rabbit does will be exposed to 0, 0.1, 1, 10, or 100 mg/kg body weight from gestation day 15 through postpartum wk 4. Reproductive sequelae in the male offspring will be determined at crucial stages of development: in frogs at 8 wk (metamorphic climax), 12 wk (when metamorphosis is complete), 24 wk (when dimorphic secondary sexual characters develop), 36 wk (when first spermatozoa are formed) and 60 wk (sexually mature), and in rabbits at 12 wk (pre-puberty), 32 wk (post-puberty), and at 100 wk (for long-term effects). The end points/parameters used to delineate the sequelae will include: evaluation of thyroid function; hypothalamic-pituitary-testicular axis; testicular descent; sexual capacity; semen characteristics; efficiency of sperm production; histopathology of testes and secondary sex organs; and immunocytochemical and ultrastructural characterization of cellular elements within the seminiferous epithelium to determine if and how germ cells manifest atypical transformation.

Expected Results:

Use of frogs and rabbits should result in development of two novel animal models. The choice of frog model will also help address another contemporary issue-reported global decline in amphibian populations-vis-a-vis ubiquitous pollutants like phthalates. Data obtained should provide a basis for establishing causal relationships, point to likely mechanisms of action, and provide preliminary baseline information for risk assessment for infantile exposures and insights into what is commonly labeled as idiopathic infertility in humans.

Publications and Presentations:

Publications have been submitted on this project: View all 16 publications for this project

Journal Articles:

Journal Articles have been submitted on this project: View all 4 journal articles for this project

Supplemental Keywords:

male reproductive development, cryptorchidism, sexual dysfunction., RFA, Health, Scientific Discipline, Toxicology, Health Risk Assessment, Chemistry, Risk Assessments, Disease & Cumulative Effects, Children's Health, Ecological Risk Assessment, Biology, risk assessment, frog deformities, dermal contact, animal model, phtalates, children, fertility, Human Health Risk Assessment, reproductive development, human exposure, children's environmental health, exposure pathways, animal studies, reproductive health, reproductive function, diopathic infertility, exposure assessment, human health risk

Progress and Final Reports:

  • 2002 Progress Report
  • 2003 Progress Report
  • 2004 Progress Report
  • 2005 Progress Report
  • Final Report